Genome-wide RNA-seq, DNA methylation and small RNA-seq analysis unraveled complex gene regulatory networks in psoriasis pathogenesis.

Psoriasis is a complex inflammatory skin disease characterized by reversible albeit relapsing red scaly plaques in the skin of a patient. In addition to the genetic predisposition, involvement of epigenetic and non-coding RNAs have also been liked with the disease. Nevertheless, any comprehensive study involving transcriptomic, small-RNA and DNA methylation at the genomic level from same patients is lacking. To investigate the complex regulation of molecular pathways in psoriasis, we carried out multi-omics integrative analysis of RNA-sequencing, small RNA-sequencing and DNA methylation profiling from the psoriatic and adjacent normal skin tissues. Our multi-omics analysis identified the genes and biological processes regulated either independently or in combination by DNA methylation and microRNAs. We identified miRNAs that specifically regulated keratinocyte hyper-proliferation, and cell cycle progression and checkpoint signaling in psoriasis. On contrary, DNA methylation was found to be more predominant in regulating immune and inflammatory responses, another causative factor in psoriasis pathogenesis. Many characteristic pathways in psoriasis e.g., Th17 cell differentiation and JAK-STAT signaling, were found to be regulated by both miRNAs and DNA methylation. We carried out functional characterization of a downregulated miRNA hsa-let-7c-5p, predicted to target upregulated genes in psoriasis involved in cell cycle processes, Th17 cell differentiation and JAK-STAT signaling pathways. Overexpression of hsa-let-7c-5p in keratinocytes caused the downregulation of its target genes, resulting in reduced cell proliferation and migration rates, demonstrating potential of miRNAs in regulating psoriasis pathogenesis. In conclusion, our findings identified distinct and shared gene-networks regulated by DNA methylation and miRNAs of a complex disease with reversible phenotype.

Serum miRNA profiling identified miRNAs associated with disease severity in psoriasis.

Psoriasis vulgaris is a chronic, autoimmune skin disease involving a complex interplay of epidermal keratinocytes, dermal fibroblast and infiltrating immune cells. Differential expressions of miRNAs are observed in psoriasis and the deregulated miRNAs are sometimes associated with disease severity. This study aims to identify miRNAs altered in the serum of psoriasis patients that are associated with the Psoriasis Area and Severity Index (PASI). In order to assess miRNA levels in the serum of psoriasis patients, we selected 24 differentially expressed miRNAs in the psoriatic skin are possibly derived from the skin and immune cells, as well as five miRNAs that are enriched in other tissues. We identified 16 miRNAs that exhibited significantly (p < 0.05) altered levels in the serum of psoriasis patients compared to healthy individuals. Among these, 13 miRNAs showed similar expression pattern in the serum of psoriasis patients as also observed in the psoriatic skin tissues. Ten miRNAs showed an accuracy of greater than 75% in classifying the psoriasis patients from healthy individuals. Further analysis of differential miRNA levels between the low PASI group and the high PASI group identified three miRNAs (miR-147b, miR-3614-5p, and miR-125a-5p) with significantly altered levels between the low severity and the high severity psoriasis patients. Our systematic investigation of skin and immune cell-derived miRNAs in the serum of psoriasis patients revealed alteration in miRNA levels to be associated with disease severity, which may help in monitoring the disease progression and therapeutic response.

A combination of circulating microRNA-375-3p and chemokines CCL11, CXCL12, and G-CSF differentiate Crohn's disease and intestinal tuberculosis.

Differentiation of Crohn's disease (CD) from intestinal tuberculosis (ITB) is a big challenge to gastroenterologists because of their indistinguishable features and insensitive diagnostic tools. A non-invasive biomarker is urgently required to distinguish ITB/CD patients particularly in India, a TB endemic region, where CD frequency is increasing rapidly due to urbanization. Among the three differentially expressed miRNAs obtained from small RNA transcriptomic profiling of ileocaecal/terminal ileal tissue of ITB/CD patients (n = 3), only two down-regulated miRNAs, miR-31-5p, and miR-215-5p showed comparable data in qRT-PCR. Out of which, only miR-215-5p was detectable in the patient's plasma, but there was no significant difference in expression between ITB/CD. On the other hand, miR-375-3p, the pulmonary TB specific marker was found in higher amount in the plasma of ITB patients than CD while reverse expression was observed in the ileocaecal/terminal ileal tissues of the same patients. Next, using Bioplex pro-human cytokine 48-plex screening panel, only three chemokines, Eotaxin-1/CCL11, SDF-1α/CXCL12, and G-CSF have noted significantly different levels in the serum of ITB/CD patients. ROC analysis has revealed that compared to a single molecule, a combination of miR-375-3p + Eotaxin-1/CCL11 + SDF-1α /CXCL12 + G-CSF showed a better AUC of 0.83, 95% CI (0.69-0.96) with 100% specificity and positive predictive value while sensitivity, negative predictive value, and accuracy were 56%, 69%, and 78% respectively in distinguishing ITB from CD. This study suggests that a combination of plasma markers shows better potential in differentiating ITB from CD than a single marker and this panel of markers may be used for clinical management of ITB/CD patients.

Co-Regulation of Protein Coding Genes by Transcription Factor and Long Non-Coding RNA in SARS-CoV-2 Infected Cells: An In Silico Analysis.

Altered expression of protein coding gene (PCG) and long non-coding RNA (lncRNA) have been identified in SARS-CoV-2 infected cells and tissues from COVID-19 patients. The functional role and mechanism (s) of transcriptional regulation of deregulated genes in COVID-19 remain largely unknown. In the present communication, reanalyzing publicly available gene expression data, we observed that 66 lncRNA and 5491 PCG were deregulated in more than one experimental condition. Combining our earlier published results and using different publicly available resources, it was observed that 72 deregulated lncRNA interacted with 3228 genes/proteins. Many targets of deregulated lncRNA could also interact with SARS-CoV-2 coded proteins, modulated by IFN treatment and identified in CRISPR screening to modulate SARS-CoV-2 infection. The majority of the deregulated lncRNA and PCG were targets of at least one of the transcription factors (TFs), interferon responsive factors (IRFs), signal transducer, and activator of transcription (STATs), NFκB, MYC, and RELA/p65. Deregulated 1069 PCG was joint targets of lncRNA and TF. These joint targets are significantly enriched with pathways relevant for SARS-CoV-2 infection indicating that joint regulation of PCG could be one of the mechanisms for deregulation. Over all this manuscript showed possible involvement of lncRNA and mechanisms of deregulation of PCG in the pathogenesis of COVID-19.

Temporal variations of country-specific mutational profile of SARS-CoV-2: effect on vaccine efficacy.

Aim: In order to curb the transmission of SARS-CoV-2, nation-wide travel restrictions at different levels were implemented in different countries. Country-specific mutational profile may exist and have an impact on vaccine efficacy. Materials & methods: We identified nonsynonymous mutations in approximately 215,000 SARS-CoV-2 sequences during the 1st year of the pandemic in 35 countries. Mutational profiles on a bimonthly basis were traced over time. We also examined the mutations that overlapped with the spike protein vaccine epitopes. Results: Several new mutations emerged over time and were dominating in specific countries. Many nonsynonymous mutations were within multiple spike protein epitopes that might impact the vaccine efficacy. Conclusion: Our study advocates requirement of active monitoring of country-specific mutations and vaccine efficacies in respective countries.

In silico analysis of altered expression of long non-coding RNA in SARS-CoV-2 infected cells and their possible regulation by STAT1, STAT3 and interferon regulatory factors.

Altered expression of long noncoding RNA (lncRNA), longer than 200 nucleotides without potential for coding protein, has been observed in diverse human diseases including viral diseases. It is largely unknown whether lncRNA would deregulate in SARS-CoV-2 infection, causing ongoing pandemic COVID-19. To identify, if lncRNA was deregulated in SARS-CoV-2 infected cells, we analyzed in silico the data in GSE147507. It was revealed that expression of 20 lncRNA like MALAT1, NEAT1 was increased and 4 lncRNA like PART1, TP53TG1 was decreased in at least two independent cell lines infected with SARS-CoV-2. Expression of NEAT1 was also increased in lungs tissue of COVID-19 patients. The deregulated lncRNA could interact with more than 2800 genes/proteins and 422 microRNAs as revealed from the database that catalogs experimentally determined interactions. Analysis with the interacting gene/protein partners of deregulated lncRNAs revealed that these genes/proteins were associated with many pathways related to viral infection, inflammation and immune functions. To find out whether these lncRNAs could be regulated by STATs and interferon regulatory factors (IRFs), we used ChIPBase v2.0 that catalogs experimentally determined binding from ChIP-seq data. It was revealed that any one of the transcription factors IRF1, IRF4, STAT1, STAT3 and STAT5A had experimentally determined binding at regions within -5kb to +1kb of the deregulated lncRNAs in at least 2 independent cell lines/conditions. Our analysis revealed that several lncRNAs could be regulated by IRF1, IRF4 STAT1 and STAT3 in response to SARS-CoV-2 infection and lncRNAs might be involved in antiviral response. However, these in silico observations are necessary to be validated experimentally.

Dysbiosis of Oral Microbiota During Oral Squamous Cell Carcinoma Development.

Infection with specific pathogens and alterations in tissue commensal microbial composition are intricately associated with the development of many human cancers. Likewise, dysbiosis of oral microbiome was also shown to play critical role in the initiation as well as progression of oral cancer. However, there are no reports portraying changes in oral microbial community in the patients of Indian subcontinent, which has the highest incidence of oral cancer per year, globally. To establish the association of bacterial dysbiosis and oral squamous cell carcinoma (OSCC) among the Indian population, malignant lesions and anatomically matched adjacent normal tissues were obtained from fifty well-differentiated OSCC patients and analyzed using 16S rRNA V3-V4 amplicon based sequencing on the MiSeq platform. Interestingly, in contrast to the previous studies, a significantly lower bacterial diversity was observed in the malignant samples as compared to the normal counterpart. Overall our study identified Prevotella, Corynebacterium, Pseudomonas, Deinococcus and Noviherbaspirillum as significantly enriched genera, whereas genera including Actinomyces, Sutterella, Stenotrophomonas, Anoxybacillus, and Serratia were notably decreased in the OSCC lesions. Moreover, we demonstrated HPV-16 but not HPV-18 was significantly associated with the OSCC development. In future, with additional validation, this panel could directly be applied into clinical diagnostic and prognostic workflows for OSCC in Indian scenario.

Characterizations of SARS-CoV-2 mutational profile, spike protein stability and viral transmission.

The recent pandemic of SARS-CoV-2 infection has affected more than 3.0 million people worldwide with more than 200 thousand reported deaths. The SARS-CoV-2 genome has the capability of gaining rapid mutations as the virus spreads. Whole-genome sequencing data offers a wide range of opportunities to study mutation dynamics. The advantage of an increasing amount of whole-genome sequence data of SARS-CoV-2 intrigued us to explore the mutation profile across the genome, to check the genome diversity, and to investigate the implications of those mutations in protein stability and viral transmission. We have identified frequently mutated residues by aligning ~660 SARS-CoV-2 genomes and validated in 10,000 datasets available in GISAID Nextstrain. We further evaluated the potential of these frequently mutated residues in protein structure stability of spike glycoprotein and their possible functional consequences in other proteins. Among the 11 genes, surface glycoprotein, nucleocapsid, ORF1ab, and ORF8 showed frequent mutations, while envelop, membrane, ORF6, ORF7a and ORF7b showed conservation in terms of amino acid substitutions. Combined analysis with the frequently mutated residues identified 20 viral variants, among which 12 specific combinations comprised more than 97% of the isolates considered for the analysis. Some of the mutations across different proteins showed co-occurrences, suggesting their structural and/or functional interaction among different SARS-COV-2 proteins, and their involvement in adaptability and viral transmission. Analysis of protein structure stability of surface glycoprotein mutants indicated the viability of specific variants and are more prone to be temporally and spatially distributed across the globe. A similar empirical analysis of other proteins indicated the existence of important functional implications of several variants. Identification of frequently mutated variants among COVID-19 patients might be useful for better clinical management, contact tracing, and containment of the disease.